Natural killer (NK) cells can be triggered to kill tumors and other cellular targets and release cytokines that regulate the immune system. However, the molecular basis for NK cell specificity and activation by cellular targets is only beginning to be appreciated. A major clue to understanding NK cell specificity was the early observation that NK cells are inhibited by the expression of MHC class I molecules on targets. This led Karre to propose the "missing-self" hypothesis in which he suggested that NK cells survey tissues for normal expression of MHC class I. In the absence of MHC class I, NK cells are released from this inhibitory influence and kill the target. In recent years, there has been an explosion of new information indicating that this inhibition is mediated by MHC class I-specific NK cell receptors that block activation. On the other hand, releasing the inhibitory influence does not automatically lead to NK cell activation. There is now abundant information indicating that the inhibitory receptors appear to regulate a second type of NK receptor that is responsible for activation. Such receptors may display their own specificity for target cell ligands but there is very little known about these receptors or their ligands. In the absence of this information, it has not been possible to fully examine the "missing-self" hypothesis and to know if the activation and inhibitory receptors are physiologically relevant and in what context. Studies on the Ly49 family of NK cell receptors, funded by the previous version of this application, have led to an appreciation for the molecular basis underlying the phenomenon of MHC class I-associated target cell resistance to mouse NK cells by illustrating the inhibitory receptor concept. In addition, the investigator's laboratory recently identified molecules belonging to the Ly49 family that can activate rather than inhibit NK cell activity. Moreover, there is now preliminary data indicating that he has identified a target displaying the putative ligand for one of these receptors. Therefore, with the availability of monoclonal antibodies specific for the Ly49 activation receptors, target cells expressing the putative ligands for one of these receptors, and genetic data indicating differential expression of an activation receptor, the specific aims are to: 1) Characterize expression and function of the activation receptors of the Ly49 family; 2) Determine ligands for Ly49 activation receptors; and 3) Examine "missing-self" hypothesis in transgenic mice expressing the ligands. Thus, these studies will provide fundamental new information that will further elucidate the function of NK cells and their tolerance mechanisms.